Automated, scaled, transposon-based production of CAR T cells

• DOI: 10.1136/jitc-2022-005189
• Publication Date: 2022-09-12T15:37:01Z
• Authors: Dominik Lock; Razieh Monjezi; Caroline Brandes; Stephan Bates; Simon Lennartz; Karin Teppert; Leon Gehrke; Rafailla Karasaka...; Teodora Lukic; Marco Schmeer; Martin Schleef; Niels Werchau; Matthias Eyrich; Mario Assenmacher; Andrew Kaiser; Sabrina Prommersb...; Thomas Schaser; Michael Hudecek
• Keywords: Transposase

Background There is an increasing demand for chimeric antigen receptor (CAR) T cell products from patients and care givers. Here, we established automated manufacturing process CAR cells on the CliniMACS Prodigy platform that scaled to provide therapeutic doses achieves gene-transfer with virus-free Sleeping Beauty (SB) transposition. Methods We used advanced connected electroporator unit performed a series of small-scale development large-scale confirmation runs primary human cells. Transposition was accomplished minicircle (MC) DNA-encoded SB100X transposase pT2 transposon encoding CD19 CAR. Results defined bi-pulse electroporation shock bi-directional unidirectional electric field, respectively, permitted efficient MC insertion maintained high frequency viable In three large scale runs, 2E8 were enriched leukapheresis product, activated, gene-engineered expanded yield up 3.5E9 total cells/1.4E9 CAR-modified within 12 days (CAR-modified cells: 28.8%±12.3%). The resulting product contained highly pure (97.3±1.6%) balanced CD4/CD8 ratio central memory phenotype (87.5%±10.4%). copy number 7.0, 9.4 6.8 in #1–3, gene analyses showed expression activation/exhaustion markers. conferred potent anti-lymphoma reactivity pre-clinical models. Notably, operator hands-on-time substantially reduced compared conventional non-automated campaigns. Conclusions report first transposon-based ready formal validation use clinical This have potential facilitate access therapy accelerate scaled, multiplexed both academic industry setting.